Division of Biological Sciences and Center for Genome Dynamics, Faculty of Science, Hokkaido University, North 10, West 8, Kita-ku, Sapporo 060-0810, Japan.
Insect Biochem Mol Biol. 2011 Aug;41(8):592-601. doi: 10.1016/j.ibmb.2011.03.011. Epub 2011 Apr 7.
Sericins are glue proteins produced specifically in the middle silk gland (MSG) of the silkworm Bombyx mori, while the silk fiber protein, fibroin, is produced in the posterior silk gland (PSG). These silk proteins are expected to be useful biomaterials in medical technology as well as biotechnology. In this study, we analyzed promoter elements of the sericin-1 gene (ser1) in vivo by introducing reporter constructs into silk glands via gene gun technology. The region from -1602 to +47 was sufficient to induce MSG-specific expression. The 5' deletion mutants showed a three-step decrease in promoter activity with the key sequences located between -1362 and -1250, -201 and -116, and -115 and -37. We detected a tissue- and stage-specific factor complex (MSG-intermolt-specific complex: MIC) bound to the sequence elements around the -1350, -320, -180, and -70 regions. A mutation in the -70 region, which inhibits MIC-binding, diminished almost all promoter activity, while another mutation that did not inhibit MIC-binding showed no effect on promoter activity. The results suggest that the binding of MIC to the above elements is intrinsic for the spatiotemporal specificity of ser1 in vivo.
丝胶蛋白是家蚕中肠丝腺特异产生的胶状蛋白，而丝素蛋白则在后丝腺产生。这些丝蛋白有望成为医学技术和生物技术中有价值的生物材料。在这项研究中，我们通过基因枪技术将报告基因构建体导入丝腺，在体内分析了丝胶蛋白-1 基因（ser1）的启动子元件。-1602 至+47 的区域足以诱导中肠丝腺特异性表达。5'缺失突变体显示启动子活性呈三步递减，关键序列位于-1362 至-1250、-201 至-116 和-115 至-37。我们检测到一个组织和阶段特异性因子复合物（中肠-蜕皮特异复合物：MIC）与-1350、-320、-180 和-70 区域周围的序列元件结合。-70 区域的突变抑制了 MIC 结合，几乎完全抑制了启动子活性，而不抑制 MIC 结合的另一个突变对启动子活性没有影响。结果表明，MIC 与上述元件的结合对于 ser1 在体内的时空特异性是内在的。
